Overall Abstract The National Resource for Translational and Developmental Proteomics, or NRTDP, proposes a constellation of activities in its second phase of operation. After launching in 2015 and establishing functional programs in each mission of a P41 BTRR, we now turn toward the proposed granting period from 2020 ? 2025 to develop new technologies and translate them by enabling both practitioners to analyze proteoforms and many collaborators to understand their biology more deeply. Consistent with changes in the P41 program, we place major emphasis on enabling basic and clinical research in proteomics, reduction to practice to extend innovations to early adopters of new implementations of top-down proteomics, which is itself a new concept to many we encounter. We also embrace mechanisms including corporate partnerships, community engagement, training and dissemination to ensure a sustainable impact even after the sunset of this P41 National Resource. After booting up our operation, we now posit three new Technology Research and Development Projects, six new Driving Biomedical Projects, a highly dynamic set of collaborative projects and an array of Community Engagement activities to continue serving our unique role within the set of P41 Resources currently supported by NIGMS. The technology and support emanating from our resource will continue to provide collaborators with orthogonal data streams and further demonstrate the value of measuring proteins more precisely than is possible with current-generation technology of ?bottom-up? proteomics. From heart disease to cancer research, neurology and age-related disorders to immunology, the analysis of proteins and proteoforms and their post- translational modifications will continue to unveil underlying molecular mechanisms through new and cell-specific technologies for precision proteomics. This will enable clinicians and cell biologists across four major disease areas, and significantly bolster American interests with an internationally-recognized excellence in a new style of mass spectrometry-based proteomics that complements existing areas of investment across the nation. The geographical distribution of collaborators and training activities are consistent with a mature resource that reaches the level of operation envisaged in the original grant proposal in 2015. The institutional support is very strong from an environment of Northwestern University on both its basic science and medical research campuses in the greater Chicago area. Leveraging a team of professional proteomicists with diverse backgrounds and experiences, the NRTDP has proven to be a highly capable enterprise that returns strongly on programmatic support received by NIGMS. As a unit, we place high value on attentiveness to the spirit and the letter of the P41 program, and also seek to synergize with other P41s within the current portfolio supported by the NIGMS and NIH.